Type I IFN induced by adenovirus serotypes 28 and 35 has multiple effects on T cell immunogenicity

Recombinant adenovirus (rAd) vectors are being investigated as vaccine delivery vehicles in preclinical and clinical studies. rAds constructed from different serotypes differ in receptor usage, tropism, and ability to activate cells, aspects of which likely contribute to their different immunogenicity profiles. In this study, we compared the infectivity and cell stimulatory capacity of recombinant adenovirus serotype 5 (rAd5), recombinant adenovirus serotype 28 (rAd28), and recombinant adenovirus serotype 35 (rAd35) in association with their respective immunogenicity profiles. We found that rAd28 and rAd35 infected and led to the in vitro maturation and activation of both human and mouse dendritic cells more efficiently compared with rAd5. In stark contrast to rAd5, rAd28 and rAd35 induced production of IFN-α and stimulated IFN-related intracellular pathways. However, the in vivo immunogenicity of rAd28 and rAd35 was significantly lower than that of rAd5. Deletion of IFN-α signaling during vaccination with rAd28 and rAd35 vectors increased the magnitude of the insert-specific T cell response to levels induced by vaccination with rAd5 vector. The negative impact of IFN-α signaling on the magnitude of the T cell response could be overcome by increasing the vaccine dose, which was also associated with greater polyfunctionality and a more favorable long-term memory phenotype of the CD8 T cell response in the presence of IFN-α signaling. Taken together, our results demonstrate that rAd-induced IFN-α production has multiple effects on T cell immunogenicity, the understanding of which should be considered in the design of rAd vaccine vectors.     

The effects of cycling cadence on the phases of joint power,crank power,force and force effectiveness

We examined the influence of cadence in cycling technique by quantifying phase relationships for a number of important variables at the crank and lower extremity joints. Any difference in the effect of cadence on force, effectiveness, and power phases would indicate an essential change in coordination pattern. Cycle kinetics was recorded for 10 male competitive cyclists at five cadences (60–100 rpm) at submaximal load (260 W). Joint powers were calculated using inverse dynamics methods. All data were expressed as a function of crank position. The phase of the crank mechanical profiles (total force, crank and joint power, and effectiveness) was calculated using four methods: crank angle of maximum (MA) and minimum (MI), fitting a sine wave (SI) and by cross-correlation (XC). These methods, apart from the MA method, showed the same relative phase. The variables, however, showed different phases being expressed as time lag: force effectiveness: 0.131 (±0.034) s; total force: 0.149 (±0.021) s; power: 0.098 (±0.027) s. The phases in joint powers hip 0.071 (±0.008), knee 0.082 (±0.009), and hip 0.077 (±0.012) were only well described by XC, and were somewhat lower than the crank power phase. These differences indicate the potential effect of inertia of the lower limb in phase shifts from joints to crank. Furthermore, the differences between the various crank variables indicate a change of technique with cadence.     

Expression and characterization of the protein Rv1399c from Mycobacterium tuberculosis. A novel carboxyl esterase structurally related to the HSL family.

The Mycobacterium tuberculosis genome contains an unusually high number of proteins involved in the metabolism of lipids belonging to the Lip family, including various nonlipolytic and lipolytic hydrolases. Driven by a structural genomic approach, we have biochemically characterized the Rv1399c gene product, LipH, previously annotated as a putative lipase. Rv1399c was overexpressed in E. coli as inclusion bodies and refolded. Rv1399c efficiently hydrolyzes soluble triacylglycerols and vinyl esters. It is inactive against emulsified substrate and its catalytic activity is strongly inhibited by the diethyl paranitrophenyl phosphate (E600). These kinetic behaviors unambiguously classify Rv1399c as a nonlipolytic rather than a lipolytic hydrolase. Sequence alignment reveals that this enzyme belongs to the alpha/beta hydrolase fold family and shares 30-40% amino acid sequence identity with members of the hormone-sensitive lipase subfamily. A model of Rv1399c derived from homologous three-dimensional structures reveals a canonical catalytic triad (Ser162, His290 and Asp260) located at the bottom of a solvent accessible pocket lined by neutral or charged residues. Based on this model, kinetic data of the Arg213Ala mutant partially explain the role of the guanidinium moiety, located close to His290, to confer an unusual low pH shift of the catalytic histidine in the wild type enzyme. Overall, these data identify Rv1399c as a new nonlipolytic hydrolase from M. tuberculosis and we thus propose to reannotate its gene product as NLH-H.     

A crucial role for the Anaplastic lymphoma kinase receptor tyrosine kinase in gut development in Drosophila melanogaster

The Drosophila melanogaster gene Anaplastic lymphoma kinase (Alk) is homologous to mammalian Alk, which encodes a member of the Alk/Ltk family of receptor tyrosine kinases (RTKs). In humans, the t(2;5) translocation, which involves the ALK locus, produces an active form of ALK, which is the causative agent in non-Hodgkin's lymphoma. The physiological function of the Alk RTK, however, is unknown. In this paper, we describe loss-of-function mutants in the Drosophila Alk gene that cause a complete failure of the development of the gut. We propose that the main function of Drosophila Alk during early embryogenesis is in visceral mesoderm development.     

Stearoyl coenzyme A desaturase 1 expression and activity are increased in the liver during iron overload

In humans, hepatic iron overload can lead to hepatocellular carcinoma development. Iron related dysregulation of hepatic genes could play a role in this phenomenon. We previously found that the carbonyl-iron overloaded mouse was a useful model to study the mechanisms involved in the development of hepatic lesions related to iron excess. The aim of the present study was to identify hepatic genes overexpressed in conditions of iron overload by using this model. A suppressive subtractive hybridization was performed between hepatic mRNAs extracted from control and 3% carbonyl-iron overloaded mice during 8 months. This methodology allowed us to identify stearoyl coenzyme A desaturase 1 (SCD1) mRNA overexpression in the liver of iron loaded mice. The corresponding enzymatic activity was also found to be significantly increased. In addition, we demonstrated that both SCD1 mRNA expression and activity were increased in another iron overload model in mice obtained by a single iron-dextran subcutaneous injection. Moreover, we found, in both models, that SCD1 mRNA was not only influenced by the quantity of iron in the liver but also by the duration of iron overload since SCD1 mRNA upregulation was not detected in earlier stages of iron overload. In addition, we found that cellular repartition likely influenced SCD1 mRNA expression. In conclusion, we demonstrated that iron excess in the liver induced both the expression of SCD1 mRNA and its corresponding enzymatic activity. The level and duration of iron overload, as well as cellular repartition of iron excess in the liver likely play a role in this induction. The fact that the expression and activity of SCD1, an enzyme adding a double bound into saturated fatty acids, are induced in two models of iron overload in mice leads to the conclusion that iron excess in the liver may enhance the biosynthesis of unsaturated fatty acids.     

Dichotomy between CD1a+ and CD83+ dendritic cells in lymph nodes during SIV infection of macaques

The prevalence and differentiation of dendritic cells (DC) in lymphoid tissue of simian immunodeficiency virus (SIV)-infected cynomolgus monkeys was studied during disease progression. Lymph node biopsies were consecutively obtained from clinical rapid and slow progressors until the development of disease consistent with simian acquired immunodeficiency syndrome (sAIDS) occurred. Quantitative evaluation of CD1a+ DC and the expression of DC antigens related to maturation (CD83, DC-LAMP and S100b) were performed at the single cell level by in situ image analysis. Despite a persistent prevalence of CD1a+ DC in lymphoid tissue during disease progression, there was a subsequent drop of mature CD83+, DC-LAMP+ and S100b+ DC, correlating with the decline of CD4+ T cells in blood. Thus, disease progression to sAIDS was associated with impaired maturation of DC, and lack of CD83, DC-LAMP and S100b expression.     

The functional profile of primary human antiviral CD8+ T cell effector activity is dictated by cognate peptide concentration

Antiviral CD8(+) T cells can elaborate at least two effector functions, cytokine production and cytotoxicity. Which effector function is elaborated can determine whether the CD8(+) T cell response is primarily inflammatory (cytokine producing) or antiviral (cytotoxic). In this study we demonstrate that cytotoxicity can be triggered at peptide concentrations 10- to 100-fold less than those required for cytokine production in primary HIV- and CMV-specific human CD8(+) T cells. Cytolytic granule exocytosis occurs at peptide concentrations insufficient to cause substantial TCR down-regulation, providing a mechanism by which a CD8(+) T cell could engage and lyse multiple target cells. TCR sequence analysis of virus-specific cells shows that individual T cell clones can degranulate or degranulate and produce cytokine depending on the Ag concentration, indicating that response heterogeneity exists within individual CD8(+) T cell clonotypes. Thus, antiviral CD8(+) T cell effector function is determined primarily by Ag concentration and is not an inherent characteristic of a virus-specific CD8(+) T cell clonotype or the virus to which the response is generated. The inherent ability of viruses to induce high or low Ag states may be the primary determinant of the cytokine vs cytolytic nature of the virus-specific CD8(+) T cell response.     

Adrenergic innervation of pancreatic islets and modulation of insulin secretion by the sympatho-adrenal system

Summary Morphological changes in the adrenergic innervation of pancreatic islets after chemical sympathectomy by use of 6-hydroxydopamine and the influence of the sympatho-adrenal system on insulin secretion were investigated in the mouse and rat.Fluorescence histochemistry revealed a clear-cut reduction in the number of adrenergic nerve fibers in the pancreatic islets 2 days after administration of 6-hydroxydopamine; the reduction was more pronounced in the rat than in the mouse. In the rat, a partial regeneration was seen after 6 weeks. In the pancreas of the mouse, after administration of 6-hydroxydopamine, a severe damage of unmyelinated nerve fibers was revealed electron microscopically. However, no ultrastructural or immunohistochemical alterations could be demonstrated in the endocrine cells of the islets.6-Hydroxydopamine induced a depression of basal plasma insulin concentrations in mice and an elevation in rats. Adrenalectomy depressed basal plasma insulin levels in mice.The -adrenoceptor antagonist phentolamine enhanced insulin secretion in normal mice. The secretory response of insulin to phentolamine was diminished by chemical sympathectomy and almost abolished by adrenalectomy or the combination of chemical sympathectomy and adrenalectomy. Thus, the effect of phentolamine is probably mediated by liberated catecholamines.It is concluded that basal insulin secretion is partially regulated by the sympatho-adrenal system and that species differences exist in this respect. In addition, the results suggest that endogenous catecholamines have the ability to promote insulin secretion.     

Immunoreactive pancreatic polypeptide (PP) occurs in the central and peripheral nervous system: Preliminary immunocytochemical observations

Summary Pancreatic polypeptide (PP) is a candidate hormone of unknown physiological significance. It is produced by a population of endocrine cells in the pancreas. In the present study a PP-like peptide was found to occur in the mammalian and avian central and peripheral nervous systems. Immunoreactive nerve fibres and nerve cell bodies were widely distributed in the brain. Dense accumulations of nerve fibres occurred in the following areas: nucleus accumbens, interstitial nucleus of the stria terminalis, para- and periventricular hypothalamic nuclei, and medial preoptic area. In addition, nerve fibres were regularly seen in cortical areas. Immunoreactive perikarya were observed in the following regions: cortex, nucleus accumbens, neostriatum and septum. In the gut, immunoreactive nerve fibers were distributed in the myenteric plexus, in smooth muscle, around blood vessels, and in the core of the villi. Immunoreactive perikarya occurred in the submucosal and myenteric plexus, suggesting that PP immunoreactive nerves are intrinsic to the gut.In the species examined, the neuronal PP-like peptide could be demonstrated with an antiserum raised against avian PP, but not with those raised against bovine or human PP. Thus, neuronal PP is distinct from the PP that occurs in pancreatic endocrine cells.